Gating of a voltage-dependent channel (colicin E1) in planar lipid bilayers: the role of protein translocation.

The voltage-dependent channel formed in planar lipid bilayers by colicin E1, or its channel-forming C-terminal fragments, is susceptible to destruction by the nonspecific protease pepsin under well-defined conditions. In particular, pepsin acts only from the cis side (the side to which colicin has been added) and only upon channels in the closed state. Channels in the open state are refractory to destruction by cis pepsin, and neither open nor closed channels are destroyed by trans pepsin. Colicin E1 channels are normally turned on by cis positive voltages and turned off by cis negative voltages. For large (greater than 80 mV) positive voltages, however, channels inactivate subsequent to opening. Associated with the inactivated state, some channels become capable of being turned on by cis negative voltages and turned off by cis positive voltages, as if the channel-forming region of the molecule has been translocated across the membrane. Consistent with this interpretation is the ability now of trans pepsin to destroy these "reversed" channels when they are closed, but not when they are open, whereas cis pepsin has no effect on them in either the open or closed state. Our results indicate that voltage gating of the E1 channel involves translocation of parts of the protein across the membrane, exposing different domains to the cis and trans solutions in the different channel states.